Single Molecule Studies of Transcription Complexes

真核生物中蛋白质编码基因的转录是一个高度调控的过程， 当无数的转录激活因子识别特定的DNA元件的近端和远端时， 核心启动子这些DNA结合的活化剂然后指导RNA的动态组装 启动子DNA上的聚合酶II、一般转录因子和辅因子复合物， 转录。由于启动转录所需的事件的确切顺序知之甚少， 对转录激活因子如何调节和指导转录的机械理解 从它们的近端和远端识别位点形成复合物对于理解它们是如何形成的至关重要。 基因表达受到调控。 为了更好地理解RNAPII介导的转录机制，我们将开发 并应用单分子技术来（i）测量GTF 和RNAPII与启动子DNA结合，（ii）确定启动子DNA结合的速率和顺序， GTF和RNAPII与启动子DNA的结合受转录激活因子的调节， 激活剂，和（iii）确定远端增强子如何激活。 在这些单分子研究中获得的关于转录起始动力学的信息将 补充所提出的生物化学，EM和X射线获得的结构信息 该PPG的晶体学研究。转录复合体形成的机制 将为抗癌治疗提供新的潜在蛋白质靶点。

Transcription of protein-encoding genes in eukaryotes is a highly regulated process that is initiated when a myriad of transcriptional activators recognize specific DNA elements proximal and distal to core promoters. These DNA bound activators then direct the dynamic assembly of RNA polymerase II, general transcription factors, and cofactor complexes on promoter DNA to initiate transcription. As the exact sequence of events needed to initiate transcription is poorly understood, a mechanistic understanding of how transcription activators regulate and direct transcription complex formation from their proximal and distant recognition sites is crucial to understanding how genes expression is regulated. Toward a better understanding of the mechanism of RNAPII mediated transcription we will develop and apply single molecule techniques to (i) measure the rates and the sequence by which the GTFs and RNAPII bind to promoter DNA, (ii) determine how the rates and the sequence by which the GTFs and RNAPII bind to promoter DNA are modulated by transcriptional activators and co- activators, and (iii) determine how activation occurs by distal enhancers. Information on dynamics of transcription initiation obtained in these single-molecule studies will complement the structural information obtained by the proposed biochemical, EM, and x-ray crystallographic studies of this PPG. Mechanistic understanding of transcription complex formation will provide new potential protein targets for anticancer therapies.